The hemoglobin types of mouse strains can be distinguished according to patterns observed on cellulose acetate electrophoresis. The two common mouse hemoglobin patterns are single and diffuse. The differences in the patterns result from differences in the β-globin chains of the hemoglobin molecules. Mice with the single hemoglobin pattern have one β-globin type identified as β-single (Hbb^s), whereas mice with the diffuse hemoglobin pattern have two different β-globin types identified as β-major (Hbb^maj) and β-minor (Hbb^min). We examined the functional and stability properties in these mouse hemoglobins, and the oxygen binding properties of red blood cells obtained from mice with four different hemoglobin phenotypes: Hbb^s/Hbb^s, Hbb^s/Hbb^min, Hbb^min/Hbb^min and Hbb^maj/Hbb^min. The P₅₀, the partial pressure of oxygen at which hemoglobin is half-saturated, of purified forms of Hbb^s, Hbb^min and Hbb^maj was 14.8 ± 0.4 mm Hg, 13.3 ± 0.6 mm Hg and 13.6 ± 0.5 mm Hg, respectively. The n value, determined from the slope of the Hill plot was 2.45 to 2.59 for the mouse hemoglobins. The alkaline Bohr effects of purified Hbb^S, Hbb^min and Hbb^maj were 0.69, 0.61 and 0.60, respectively. The mechanical stability of Hbb^s, Hbb^min and Hbb^maj, expressed by the first order kinetic constant, were 0.098 ± 0.01/min, 0.027 ± 0.013/min and 0.27/min, respectively. The P₅₀ of red blood cell suspensions from lines of mice expressing Hbb^s/Hbb^s, Hbb^min/Hbb^min, Hbb^s/Hbb^min and Hbb^maj/Hbb^min were 40.2 ± 1.8 mm Hg, 40.4 ± 1.5 mm Hg, 38.9 ± 1.4 mm Hg, and 38.7 ± 0.9 mm Hg, respectively.